Generation of hydrogen from hydrocarbon bearing materials

ABSTRACT

Disclosed are strategies for the economical microbial generation of hydrogen, useful as an alternative energy source, from hydrocarbon-rich deposits such as coal, oil and/or gas formations, oil shale, bitumen, tar sands, carbonaceous shale, peat deposits and sediments rich in organic matter through the management of the metabolism of microbial consortia.

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is a continuation, and claims the benefit, ofco-pending, commonly assigned U.S. patent application Ser. No.13/310,461, filed Dec. 2, 2011, entitled “Generation of Hydrogen FromHydrocarbon Bearing Materials,” which is a continuation of U.S. patentapplication Ser. No. 11/568,974, filed Jun. 22, 2007, entitled“Generation of Hydrogen From Hydrocarbon Bearing Materials,” which is aU.S. National Phase Application of PCT/US05/16124, filed May 6, 2005,entitled “Generation of Hydrogen From Hydrocarbon Bearing Materials,”which claims the benefit of U.S. Provisional Application No. 60/570,174,filed May 12, 2004, entitled “Generation of Hydrogen From HydrocarbonBearing Materials.” The entire disclosures of all of which areincorporated herein by reference for all purposes.

BACKGROUND OF THE INVENTION

The field of the present invention is microbial production of hydrogenthrough the management of the anaerobic or nearly anaerobic metabolismof consortia of microorganisms, including archaea and bacteria, toproduce hydrogen, either in-situ or ex-situ, from hydrocarbon substratessuch as coal, carbonaceous shale, oil, tar sands, bitumen, peat, and thelike.

Because of the clean burning nature of hydrogen arid its energy densityon a weight basis, it is highly valued as an energy source. Billions ofdollars of research have been expended on the invention and refinementof hydrogen fuel cells which have none of the carbon emissionsassociated with the use of fossil fuels. The greatest single obstacle tothe widespread use of hydrogen fuel cells for motor transport andelectricity generation is the high cost of molecular hydrogen on a costper Btu basis relative to gasoline, coal, and natural gas. The presentinvention can dramatically lower the cost of hydrogen by utilizingmicrobial consortia to generate that hydrogen from the vast resources ofcoal, carbonaceous shales, oil, tar sands, bitumen and peat availablethroughout the world.

Currently, hydrogen is generated primarily by reformulation of methaneby exposure to high pressure, high temperature steam. Most of thehydrogen liberated in this reaction is used in combination with nitrogento make fertilizer. However, the Btu value of the hydrogen produced isfar less than the Btu value of the fuel needed to produce it, makingthis an expensive and endergonic reaction that frustrates the widespreaduse of hydrogen as a transportation or electrical generation fuel.

Unlike the substrates in the present invention, agricultural waste,compost, municipal wastes including sewage and waste waters have beenutilized as starting materials for fermentations to yield hydrogen gas.For example, Clostridia have been identified as important microorganismsfor the microbial production of hydrogen gas from agricultural wastes,other cellulosic materials and sewage (JP 07031998 (1995), Van Ginkel etal. (2000) Ann. Conf. & Expos. on Water Quality and WastewaterTreatment, Water Environment Federation, 3413-3429; Nazinia, Tenn.(1981) Mikrobiologlia 50: 163-166). A wide variety of heterotrophicmicroorganisms are known to produce hydrogen gas from organic wasteproducts. Photosynthetic microorganisms such as species of Rhodobacter,Rhodopseudomonas and Rhodospirillum have also been proposed asmicroorganisms useful in the microbial production of hydrogen. Apotential disadvantage of using photosynthetic microorganisms forphototrophic generation of hydrogen relates to limitations imposed onthe penetration of light into a reactor containing a relevant substrateor where the substrate is underground, such as in massive sub-bituminouscoal deposits containing billions of tons of coal and billions of poundsof hydrogen.

Despite the advances that have been made towards producing hydrogen fromagricultural and municipal wastes, the limited availability andinconsistent composition and quality of these materials as well as thecost of hydrogen production through this process have precluded theiruse to date as substrates for the microbial production of hydrogen atquantities that could create a significant alternative source of energyrelative to traditional fossil fuels (i.e. crude oil, natural gas, andcoal). Furthermore, even assuming a substantial increase in biomassdevoted to H₂ production, maximum total H₂ generated from agriculturaland municipal wastes can provide less than 5% of current U.S. energydemand.

Research on microbial hydrogen production has focused exclusively on theconversion of waste products that contain easily fermentable materialsincluding polysaccharides (Wang et al. 2003). For example, a myriad ofcellulose-containing wastes produced in the food processing industry,agriculture, and domestic sewage have been evaluated as substrates tosupport microbial hydrogen production. Cellulose and otherpolysaccharides are easily fermentable (to hydrogen, carbon dioxide,acetate, and other organic acids) by a number of well characterizedmicroorganisms and metabolic pathways. However, arguments have been madethat microbial hydrogen production using these substrates will requirevery high conversion rates and efficiencies that have not been attainedby the tested microorganisms (see, e.g. Benemann (1996)).

Even though there have been substantial technological advances in fossilfuel production techniques, the majority of oil discovered in the worldremains trapped in the subsurface. Trapped crude oil in oil reservoirs,coals that may be too deep to excavate or that contain levels ofimpurities too high to burn, and carbonaceous shales that provide only asmall amount of natural gas relative to the total hydrogen and energywithin them represent a large source of substrate for microbialconversion to hydrogen.

There are numerous oil fields within the United States and around theworld that are at or near the point of abandonment due to the inabilityto continue to produce oil from them profitably. Under currenttechnology and oil prices, those fields will be abandoned with billionsof barrels of oil remaining in place since primary and secondary oilrecovery techniques still normally leave behind half or more of theoriginal oil in place in those reservoirs at the time they areabandoned. That remaining oil represents a significant quantity ofsubstrate for the generation of hydrogen that would otherwise be lost.Hydrocarbon-bearing formations have been noted to contain variableamounts of hydrogen gas. See, e.g., Khorunzhii et al. (1977)Ugol'Ukrainy 4:42-44; Kosenko et al. (1967) Geologichnii Zhurnal27:83-87. Although there is apparent recognition of the presence of somehydrogen in coal formations, it is unusual to detect hydrogen inhydrocarbon deposits. The present inventors are not aware of reportsthat document the microbial production of hydrogen from these materialsor of commercial operations in which hydrogen is produced by microbialmetabolism in coal-bearing or other hydrocarbon-rich environments.

By managing the metabolism of microorganisms to generate hydrogen, largeamounts of that clean fuel can be made available for use. The substratefor that hydrogen generation is available in vast quantities in the formof coal, carbonaceous shale, tar sands, bitumen, peat, and the remainingoil in underground reservoirs.

BRIEF SUMMARY OF THE INVENTION

The term “hydrogen” is used herein to denote both molecular hydrogen(H₂) and atomic hydrogen (H).

The present inventors have discovered that at least a portion of thehydrogen detectable in hydrocarbon deposits (coal, bitumen, oil shalecarbonaceous shale, tar sands, peat, oil and/or gas formations,sediments rich in organic matter and the like) is produced by microbialmetabolism. Furthermore, the inventors have discovered that in certainhydrocarbon formations the hydrogen is a major precursor for biogenicmethane production. The present inventors have demonstrated thatbiosynthetic hydrogen generation can be enhanced by interventions thatstimulate microorganism growth and metabolism in hydrocarbon formations.

It is an object of the present invention to provide a method forproducing hydrogen through the management of the metabolism ofmicroorganisms acting on hydrocarbons in natural materials, such ascoal, bitumen, oil shale, carbonaceous shale, tar sands, peat, oiland/or gas formations, sediments rich in organic matter, and the like.This invention can be utilized either to produce that hydrogen as afree, molecular hydrogen end-product through the inhibition of its useby hydrogen consumers, such as methanogenic microorganisms, in thehydrocarbon substrate and the recovery of that free molecular hydrogen,or to allow the free hydrogen to be consumed by methanogenicmicroorganisms and recover that hydrogen as part of the methane oracetate produced by microorganisms. Hydrogen production can occurin-situ or it can occur ex-situ, for example, in a bioreactor, afterremoval of the hydrocarbon-rich material from the geologic formation inwhich it is found. The microbial metabolism can be achieved by themicroflora which occurs naturally within the formation or it can be theresult of metabolic reactions of one or more microorganisms introducedinto the hydrocarbon rich geologic formation or bioreactor in which thehydrogen is produced.

In hydrocarbon deposits studied to date, the inventors have determinedthat H₂ availability is rate-limiting for biogenic methane productionwithin the deposit. Enhancement of microbial hydrogen production cantherefore lead to enhanced methanogenesis in the deposit, where that isthe desired product.

The present inventors have determined that the production of hydrogenfrom hydrocarbon rich substrates, preferably in anaerobic or nearlyanaerobic conditions, can be enhanced by several fold over naturalmicrobial hydrogen production through the management of the metabolismof microorganisms that exist within or can be introduced into thathydrocarbon substrate, whether in-situ or ex-situ. The management of theconsortia of microorganisms is achieved through selective introductionof nutrients, by other amendments to the substrate and the fluidscontained therein, or through the introduction of new or alteration ofexisting microorganisms into the microbial consortia which are capableof producing hydrogen.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a schematic diagram of the anaerobic microbial metabolicpathways detected in the tested coal seam.

FIG. 2 is a bar graph showing total hydrogen processed, includinghydrogen gas converted to methane and acetic acid as well as accumulatedhydrogen gas, in sub bituminous coal (from the Dietz coal seam) slurrieswith various amendments as described.

FIG. 3 shows the hydrogen gas accumulated during incubation of coalslurries (from the Dietz coal seam) with various amendments asdescribed.

FIG. 4 is a graph showing increased methane production as a function ofincreased hydrogen production.

DETAILED DESCRIPTION OF THE INVENTION

In a particular embodiment, the environmental parameters of thehydrocarbon-rich materials are modified so as to improve production ofhydrogen, for example by decreasing salinity or by introducing anutrient (metal ion(s), nitrogenous compound(s), phosphorus-containingcompound(s), vitamin(s), complex nutrients, or the like) which islimiting in the formation environment or in the reactor containing thehydrocarbon-rich natural material. Environmental factors which can bemanipulated in the formation include, without limitation, temperature,salinity, sulfate content, water content, pH, nitrogen source andamount, phosphorous source and amount, trace elements and vitamins. Itcan be beneficial to measure the content of one or more chemicalcomponents of the formation or of the water within the formation in aneffort to identify specific nutrient limitations to allow formulation ofsupplements useful to improve the rate and/or extent of hydrogenformation. The hydrogen which is evolved as the result of microbialmetabolism is captured using equipment known to the art, such as thatused for the capture of natural gas or methane. Where the gaseousmaterial produced in the hydrocarbon rich geologic formation in thein-situ application of the invention or in a bioreactor in the case ofex-situ application of the invention is a mixture of gases, for example,consisting of carbon dioxide, methane and hydrogen, readily availableseparation technology can be employed so that one or more gases ofinterest can be collected. In, addition, or in the alternative, one ormore inhibitors of hydrogen-consuming microbial processes, includingmethanogenesis, acetogenesis or sulfate reduction, can be introducedinto the hydrocarbon rich geologic formation or the bioreactor in anamount such that free molecular hydrogen accumulation is enhancedbecause of the inability of hydrogen consuming microorganisms to utilizethat hydrogen due to the inhibitors introduced to prevent theirmetabolic consumption of that hydrogen.

Inhibitors of methanogenesis include for example, without limitation,2-bromethanesulfonate (BESA) and certain derivatives of p-aminobenzoicacid. Inhibitors of acetogenesis include, for example, withoutlimitation, monensin and tetracycline. In addition, the environmentalparameters in the formation (or bioreactor) can be manipulated so as toselectively inhibit methanogens and/or other hydrogen-consumingmicroorganisms. Sulfate reduction by microorganisms and the concomitantconsumption of hydrogen in that reaction is inhibited by molybdate. Theaddition of inhibitors is not required in environments with conditionsthat do not favor the terminal hydrogen-consuming reactions.

As specifically exemplified, for Powder River Basin coal core samples,hydrogen production is optimized through the amendment of the samplewith an aqueous solution including metal ions. Desirably, theconcentration of each metal ion (copper, iron, nickel and cobalt) in thesolution is from about 0.005 to 10 mg/liter. Alternatively, or inaddition, the solution can comprise minerals, including ammoniumphosphate and/or additional compounds, and it can further, oralternatively, comprise a complex nutrient such as yeast extract (YE) orinexpensive waste products containing similar complex nutrients such asthose from fermentation plants, distilleries, breweries or bakeries,among others.

The microbial composition of the formation can itself be modified. Itcan be advantageous to analyze the geologic formation or thehydrocarbon-rich material from which hydrogen is to be produced todetermine the presence of one or more particular microorganisms. In atleast some instances, the nutritional and/or environmental preferencesof the microorganism(s) which directly or indirectly effect hydrogenproduction are known, and by comparing the physical characteristics ofthe formation or hydrocarbon rich material with the preferences of atleast one microorganism effecting hydrogen production, one recognizesthe amendments to the formation or bioreactor which increase the rate orfinal yield of hydrogen production.

The anaerobic decomposition of complex sources of organic matter,including hydrocarbons, is catalyzed by microbial consortia. In general,the initial stages of decomposition include microbial depolymerizationreactions and/or the production of organic acids, alcohols, hydrogen,and carbon dioxide. The organic acids and alcohols are furthermetabolized by other microorganisms to acetic acid, hydrogen, and carbondioxide. The produced hydrogen and acetic acid are the primarysubstrates for the terminal members of the “anaerobic food chain”including methanogenic archaea, sulfate-reducing bacteria, andmicroorganisms that use oxidized metals, including ferric iron, aselectron acceptors. Therefore, the anaerobic decomposition of organicmatter is dependent on the interaction between these “cross feeding”functional groups within the consortia.

The importance of this “cross feeding” and of anaerobic microbialconsortia in general, is perhaps best demonstrated by the finding thatthe bioconversion of many organic acids is often catalyzed viasyntrophic metabolism. For example, in some ecosystems, many of theorganic acids are biodegraded by syntrophic microorganisms that aredependent on the terminal members of the food chain (includingmethanogens and methanotrophs) to maintain hydrogen and/or acetate atlow concentrations. Removal of particular metabolic intermediates andproducts, including hydrogen, either by collection or by furthermetabolism, can therefore enhance the conversion to hydrogen of complexhydrocarbons such as coal, oil, carbonaceous shales, bitumen, tar sands,and peat. The interactions between various functional groups of theconsortia, and between the anaerobic or nearly anaerobic microbialconsortia with the local environmental conditions in the formation, havea strong impact on the predominant metabolic pathways and the gaseousend-products produced (Le. hydrogen, methane and/or carbon dioxide).Similarly, modification of one or more environmental parameters of theformation can influence metabolic activity.

In addition, the microbial populations within the formation can also bemanipulated; one or more particular organisms or a microbial consortiumor microbial consortia can be introduced to increase the rate ofmetabolism of the hydrocarbons to molecular hydrogen within theformation. The introduced microorganism(s) can have a genetic contentwhich occurs in nature, or the genetic content of the microorganism(s)can be modified in the laboratory; i.e., a naturally occurring plasmid,transposon or other nucleic acid molecule can be introduced to optimizean existing metabolic capability or empower the microorganism ormicroorganisms to carry out a metabolic reaction which it or they hadnot been capable of previously. Alternatively, the introduced nucleicacid molecule can be one which has been created in the laboratory tooptimize an existing metabolic capability or to encode one or moreproteins which allow the microorganism or microorganisms into which itis introduced to carry out a metabolic reaction which it or they couldnot carry out in nature. Alternatively, the plasmid, transposon or othernucleic acid molecule introduced into a microorganism of interest can becreated by joining portions of genetic material in the laboratory whichdo not occur joined in nature. For example, without wishing to be boundby any particular theory, the present inventors believe that benzoatemetabolism is a bottleneck in the breakdown of complex hydrocarbonmolecules to among other things, molecular hydrogen, in at least somehydrocarbon rich formations. The specific enzymatic reactions within thebenzoate-degrading microorganisms that limit benzoate metabolism can betargeted for genetic alteration. The nucleic acid molecule of interestcan be introduced by any of a variety of ways known to the art,including but not limited to transformation, transfection, conjugationor transduction. In addition, the genetic content of a microbe ofinterest can be altered by selecting or screening for a novel phenotypictrait of interest, as is well known to the art. While photoevolution ofhydrogen is not likely to be practical in the context of hydrogenproduction from hydrocarbon deposits, certain of the phototrophicbacteria can evolve hydrogen through the action of formate hydrogenlyaseduring anaerobic, dark fermentation of organic substrates such aspyruvate (e.g., Rhodospirillum rubrum, see Drews and Imhoff (1991) inVariations in Autotrophic Life, Academic Press, NY, pp. 51-97).Rhodopseudomonas palustris, Rhodobacter capsulatus, Rhodopseudomonasmolischianum and Chromatium minutissimum are nonlimiting examples ofmicroorganisms that produce hydrogen with suitable carbon sources, suchas glucose and formate, in the dark. Other electron donors for themicrobial production of hydrogen (in the dark) can include, withoutlimitation, organic acids, alcohols, amino acids and carbohydrates,depending on the microorganism or combination of microorganisms. For areview, see Sasikala et al. (1993) Adv. Appl. Microbiol. 38:211-295.

Environmental parameters within a formation can be assessed by obtainingeither a solid phase sample such as a core sample, drill cutting and/ora sample of water, or gaseous or liquid samples, from within theformation. Appropriate parameters to assess in core samples include thepresence and population of hydrogen-producing microorganisms and theiractivity, sulfate reducing microorganisms and their activity,methanogens and methanogenic activity, acetogens and acetogenicactivity, gases including hydrogen, carbon dioxide, methane, and carbonmonoxide, acetate and other organic acids, salinity, cation content,concentrations of copper, cobalt, zinc, iron, nickel, nitrogenouscompounds, and phosphorus-containing compounds, sulfate content, watercontent and the content and the nature and amounts of carbonaceousmaterials. For aqueous samples taken from the formation, it is useful tomeasure one or more parameters including pH, salinity, concentrations ofsulfate, sulfide, and acetic acid, metal content, nitrogen content andphosphorous content, among others. Information from the assessments canbe used to determine optional supplementation of water ornutrient-containing compositions which can be pumped into the formationto improve hydrogen generation.

Core samples, preferably taken and maintained under anaerobicconditions, can be used to estimate the intrinsic rate of hydrogen (orother product) generation with or without supplementation with anynutritional parameter or inhibitor of interest. Desirably, surfacemicroorganisms are not introduced as contaminants in the sample prior toculture experiments in the laboratory. In such cases, the sample is keptin a sealed environment. Methodologies for gas sampling and analysis arewell known to the art.

By implementation of the present invention, the economic lives ofabandoned oil fields and biogenic natural gas projects can be extendedindefinitely by means of utilizing the existing infrastructure such asdrilled and cased wells in those fields to access and manipulate thosereservoirs as described herein to stimulate hydrogen production fromthem and to produce that hydrogen for beneficial use. Generatinghydrogen in such a manner not only produces a clean fuel for use inmotor transport and electricity generation, but it also reduces the needto drill new wells in potentially environmentally sensitive areas toaccess more natural gas. By utilizing the existing infrastructure inexisting fields, the present invention can potentially save billions ofdollars in capital expenditures while simultaneously generating avaluable energy source and minimizing the number of wells to be drilledto access that energy source.

Microbial consortia, either metabolically active or dormant, are presentin environments which are rich in hydrocarbons, including deposits ofcoal, bitumen, oil, tar sands, peat, oil shales, natural gas deposits,carbonaceous shales, and sediments rich in organic matter. As usedherein, hydrocarbons means compounds comprising at least hydrogen andcarbon. In the context of this application, the term “microorganisms” isintended to encompass the bacteria and the archaea. Microorganisms cansurvive in and/or multiply in a wide range of environments,characterized by vast variations in temperature, pressure, pH, oxygenconcentration, substrate availability and salinity. It is noted that notall microorganism strains can tolerate the entire ranges ofenvironmental parameters. Thus, the present methods improve themetabolism of the consortia in hydrocarbon-rich environments, forexample, in coal deposits, by altering reactor conditions or the in-situgeologic formation environment so that the microbial production ofhydrogen gas (or other metabolic product(s) of interest) is maximized.

Methanogenesis is a metabolic process that commonly occurs in anaerobicsediments containing decomposing organic matter. Methane is oftenpresent within hydrocarbon-rich formations as adsorbed (onto solids),free, and dissolved methane. Some of the methane is “old” methaneproduced by methanogenesis upon sediment deposition or thermochemicallyupon sediment buried to significant depths; in many formations some ofthe methane is “new”, i.e. it is newly synthesized via recent or ongoingmethanogenesis. The present inventors have confirmed by use ofradiotracers that microorganisms in all the coal samples tested to dateare metabolically active. Methanogenesis, supported by the availabilityof molecular hydrogen to methanogenic microorganisms that consume thatmolecular hydrogen, has been detected in all coal samples that are freeof sulfate, the presence of which has been seen to coincide with a lackof methane. Field geochemical signatures, attest to the occurrence ofmethanogenesis and hydrogen turnover in situ. That is, carbon andhydrogen isotope patterns of methane, carbon dioxide, and formationwater also indicate the occurrence of methanogenesis supported by thepresence of molecular hydrogen that is consumed by methanogens andacetogens.

The current invention utilizes complex and more recalcitrant hydrocarbonsubstrates in shales, coals, oils, and other carbonaceous materials assubstrates for microbial hydrogen production. Prior to the currentinvention, these materials had not been tested as substrates formicrobial hydrogen production. In contrast to cellulosic materials, thebiodegradation of hydrocarbons is likely dependent on anaerobicmicrobial consortia. Specific microorganisms, metabolic pathways, andinteractions between members of the consortia are not fullycharacterized. Results presented herein indicate that coal can bebiodegraded via microbial consortia indigenous to the coal, and atsignificant rates once nutritional limitations are overcome. As opposedto widely dispersed cellulosic wastes, vast quantities of hydrocarbonwithin localized deposits are available as substrates for anaerobicmicrobial hydrogen production. For example, large quantities of coal atmines or coal fired power plants could be sites for ex-situ biosynthesisof hydrogen. In-situ application can occur within natural gas productionfields (including oil, coal bed methane, and shales) with existing wellsand transport facilities that can be converted for hydrogen productionand distribution. Clearly, lower hydrogen conversion rates andefficiencies are required where larger quantities of substrate andexisting infrastructure are available.

Although it is possible to introduce exogenous microorganisms withmetabolic capabilities of interest, it is preferred to stimulatemetabolic activities of the indigenous microorganisms. Limitations onmicrobial generation of a gas of interest, especially hydrogen but alsopotentially methane, include, but are not limited to, formationtemperature, the nature and distribution of microorganisms within thehydrocarbon-rich deposit or formation, nutrient availability within thedeposit (including vitamins, nitrogen, phosphorus, trace metals such asnickel, cobalt, iron and copper), the pH, availability of competingacceptors of reducing power such as sulfate or ferrous iron, and a lackof anaerobic conditions, among others. It is believed that it is easierto provide nutritional supplements as aqueous solutions or aerosols orwater to the indigenous microorganisms than to transport microorganismsthrough a geologic formation, and the use of solutions of interest isless costly than maintaining and growing microorganisms at or near theformation. Water content and movement are also important factors fromthe standpoint of supporting microbial life and nutrient transport,respectively. An important physical factor is the permeability and/orfractures (natural or manmade) within the geologic formation as thisaffects nutrient and gas transport within the formation environment andaffects collection of a gaseous products of interest such as hydrogen,and also potentially methane. Movement of introduced materials throughthe formation can be improved by fracturing and/or horizontal drilling.

We have demonstrated, in laboratory scale experiments, hydrogen gasaccumulation in anaerobic coal slurries to approximately an order ofmagnitude greater than concentrations typically observed in anaerobicorganic-rich environments where the rate of microbial hydrogenproduction is generally balanced by hydrogen consumption. The productionof hydrogen in the coal slurries was proven to be effected by ananaerobic microbial consortium or consortia indigenous to the coal.Hydrogen gas accumulated to the greatest extent when coal biodegradationwas stimulated by overcoming nutrient limitations, coupled withinhibiting methanogenesis, which was the predominant terminal electronaccepting reaction and one of the hydrogen-consuming reactions in thecoal samples. These results are believed to demonstrate for the firsttime the generation of hydrogen gas from coal using indigenousmicroorganisms. Other similar hydrocarbon-rich materials can also serveas substrates for microbial hydrogen production.

Methods and equipment for collection of gases from a formation are wellknown to the art. See, e.g., WO 02/34931 for a discussion. Strategiesare also well known for optimum well placement in a geologic formation.Similarly, methods and equipment for collection of gaseous products froma bioreactor are also well known in the art. See, for example, U.S. Pat.No. 6,340,581.

All references cited in the present application are incorporated byreference herein to the extent that there is no inconsistency with thepresent disclosure.

The following examples are provided for illustrative purposes, and arenot intended to limit the scope of the invention as claimed herein. Anyvariations in the exemplified articles that occur to the skilled artisanare intended to fall within the scope of the present invention.

EXAMPLES Example 1 Core Samples

Core samples were obtained from regions of interest within coal beds. Tominimize opportunities for microbial and/or chemical contamination ofcoal samples, no drilling muds were used during sample collection. Thecenters of core samples were desirably used for laboratory studies tofurther minimize the potential for contamination. The core samples werestored and transported in sealed stainless steel canisters which hadbeen purged with argon to maintain an anaerobic environment so thatobligately anaerobic microorganisms were not lost. In most cases, themajority of adsorbed gases were removed from the core samples prior toanalysis or culture experiments.

Core samples from the Tongue River area in the northwest portion of thePowder River Basin, Dietz coal seam, were utilized for theseexperiments. The core was collected in the field in a manner thatprevented oxygen exposure by placing it into a steel canister that waspurged with argon. Gas in the core used for the experiments describedherein was extensively desorbed during long-term storage and previoususe.

Coal samples were passed through a small jaw crusher twice resulting inpulverized coal (to yield a particle size similar to that of granulatedsugar) that was measured into serum bottles (5 g per bottle);manipulations were carried out in an anaerobic environment. Anoxicformation water that was collected from wells in the Dietz coal seam wasadded to the coal samples. Sodium sulfide (to 0.5 mM) was added to thewater samples to ensure strict anoxic conditions in the coal slurriesduring long-term incubation. The bottles containing the coal slurrieswere sealed with butyl rubber stoppers and purged with helium, resultingin a small overpressure (˜3 psi). Radioactively labeled ¹⁴C-bicarbonatewas added to the incubations to trace methanogenic activity fromhydrogen (4H₂+¹⁴CO₂→¹⁴CH₂+2H₂O) in an effort to confirm the microbialproduction of methane from hydrogen gas.

Example 2 Nutritional Supplements

Nutritional supplements for stimulating coal biodegradation and hydrogenproduction were added to the coal slurries from anoxic stock solutions.Most of the nutritional supplements tested are inorganic compoundsneeded in very low or trace concentrations. These include a nitrogensource as ammonia, a phosphorous source as phosphate, a mixture ofammonia and phosphate, a mineral solution that contains phosphate,ammonia, and potassium, and yeast extract which contains a complexmixture of inorganic nutrients and organic precursors for cell growth.The compositions of the Mineral, Vitamin, and Trace Metal Solutions usedin the experiments are listed in Tables 1-3. The Mineral Solution is a200× concentrate, the Vitamin Solution is a 500× concentrate, and theTrace Metal Solution is a 100× concentrate.

TABLE 1 Mineral solution^(a) Component Amt (g)/liter Sodium chloride 80Ammonium chloride 100 Potassium chloride 10 Potassium phosphate 10Magnesium Sulfate x 7H₂O 20 Calcium chloride x 2H₂O 4 ^(a)A solutioncontaining the major inorganic components required for microbial growth

TABLE 2 Vitamin solution^(a) Component Amt (g)/liter Pyridoxine-HCl 10Thiamine-HCl 5 Riboflavine 5 Calcium pantothenate 5 Thioctic acid 5p-Aminobenzoic acid 5 Nicotinic acid 5 Vitamin B₁₂ 5 MESA^(b) 5 Biotin 2Folic acid 2 ^(a)A solution designed to meet water-soluble vitaminrequirements of many microorganisms ^(b)Mercaptoethanesulfonic acid

TABLE 3 Trace metal solution^(a) Component Amt (g)/literNitrilotriacetic acid 2.0 Adjust pH to 6 with KOH Manganese sulfide 1.0Ferrous ammonium sulfate 0.8 Cobalt chloride 0.2 Zinc sulfate 0.02Copper chloride 0.02 Nickel chloride 0.02 Sodium molybdate 0.02 Sodiumselenate 0.02 Sodium tungstate 0.02 ^(a)A solution designed to meet thetrace metal requirements of many microorganisms.

Sterile samples were autoclaved (1200 C for 20 minutes) on threeoccasions.

Bromoethanesulfonic acid (BESA), a methanogen inhibitor, was added toselected incubations. Coal slurries without nutritional supplements orinhibitors were also prepared. Hydrogen production in the amendedslurries relative to sterile coal slurries was measured to identify thedegree of stimulation.

After approximately 134 days of incubation, all of the coal slurrieswere sampled for the aqueous phase (0.5 of 10 ml) and for gas analysisincluding hydrogen, methane, carbon dioxide, radioactive methane, andradioactive carbon dioxide. Methane and carbon dioxide were determinedusing a Hewlett Packard 5890 gas chromatograph equipped with a thermalconductivity detector. Radiolabeled methane and carbon dioxide weredetermined using a gas proportional counter (Innus Systems, Tampa,Fla.). Results are reported herein as mL hydrogen gas produced per kg ofcoal calculated on a per year basis.

The total quantity of hydrogen that was produced and consumed in theDietz coal slurries was estimated as the sum of hydrogen that wasconsumed during methanogenesis to make methane, used by acetogenicmicroorganisms to make acetate, and the hydrogen that accumulated in theheadspace of the incubations. The stoichiometric relationships of thehydrogen consuming processes identified in the coal samples with respectto hydrogen are:

4H₂+CO₂→CH₄+2H₂O

4H₂+CO₂→CH₃COOH(acetate)+2H₂O.

Therefore, the calculation to estimate the total 1-1 mol of hydrogenthat was processed (produced and consumed) is: (μmmol CH₄×4)+(μmmolacetate×4). This value was added to the quantity of hydrogen (in μmmol)that accumulated in the headspace to obtain the total hydrogen values.FIG. 2 illustrates the average calculated total hydrogen processed withvalues shown (converted to units of mL H₂/kg/year) for triplicate Dietzcoal slurries with a variety of treatments. All samples were analyzedtogether within 36 hours in order to minimize correction required forbarometric pressure, instrumental variations and the like. Very littleactivity was observed in the sterile controls, thereby proving that theprocessed hydrogen is of biogenic origin. The highest total hydrogenvalue (about 4121 mL H₂/kg coal/yr; or about 0.6 lbs H₂/ton coal/yr) wasobtained in incubations that were amended with a mixture of minerals andvitamins. Incubations amended with various nutrient supplements incombination with yeast extract also processed significantly higherquantities of hydrogen relative to the sterile and unamended (nonutrient supplements in addition to formation water) controls. Coalslurries that were treated with the methanogenic inhibitor BESA incombination with a minerals mixture, metals mixture, vitamins, and yeastextract, also processed significant quantities of hydrogen (primarilythrough acetate production, data not shown). This set of experimentsconfirms that significant quantities of hydrogen were processed in thesecoal slurries, especially when nutritional limitations of the microbialconsortium or consortia were overcome. The quantity of hydrogen gas thataccumulated in the headspace of the coal slurries is illustrated in FIG.3. Hydrogen concentrations in the sterile slurries were below thedetection limit (equivalent to approximately 0.003%). Hydrogenaccumulated to the greatest extent when methanogenic activity wasinhibited with BESA. For example, incubations supplemented with bothBESA and a suite of nutrient supplements (minerals, metals, vitamins,and yeast extract) accumulated hydrogen to levels approximately an orderof magnitude over the accumulation of a similarly stimulated sample(minerals and vitamins and metals and YE) without BESA. Theseexperiments demonstrate the utility of inhibiting methanogenic activity,especially in concert with nutrient supplementation, to increase theproduction and accumulation of free, molecular hydrogen.

FIG. 4 graphically demonstrates the direct relationship observed betweenbiogenic hydrogen production and methane synthesis. The data demonstratethat as hydrogen biosynthesis was increased, methane synthesis wasincreased in direct proportion. Even at the highest rate of hydrogensynthesis, the proportionality was observed, indicating that hydrogenbiosynthesis was rate-limiting for methanogenesis in the samples thatwere studied. Two points on the ordinate demonstrate the effect of BESAas inhibitor of methane biosynthesis and resultant accumulation ofhydrogen. It will be understood by those skilled in the art that inother types of hydrocarbon deposit, other formations biosynthetichydrogen may be utilized in processes other than methanogenesis, forexample in production of acetate. Since acetate is itself a producthaving economic value, stimulation of biogenic acetate production insuch formations can be of value, in addition to the value of thehydrogen and/or methane produced.

In summary, these results indicate that the anaerobic bioconversion ofthe tested coals is, at least in part, restricted by the availability ofnutrients within the formation water and coal samples used to preparethe slurries. When these nutrient limitations were alleviated, anaerobiccoal bioconversion to produce hydrogen gas as an end product wasstimulated or if methane is the targeted end product, the formation ofmolecular hydrogen as an intermediate can stimulate methane production.Hydrogen gas accumulation can be improved by inhibiting methanogenesis.Other hydrocarbon-rich substrates or formations including, but notlimited to, oil and natural gas deposits, peat, bitumen, tar sands,carbonaceous shale and sediments rich in organic matter can be similarlytreated to improve hydrogen production and/or accumulation.

What is claimed is:
 1. A method of enhancing microbial production ofmethane from a hydrocarbon-rich deposit, the method comprising:characterizing at least one environmental parameter for the in situhydrocarbon-rich deposit; introducing an aqueous solution to the in situhydrocarbon-rich deposit, wherein the aqueous solution stimulates amicrobial consortium to increase a production rate for producing themethane from the in situ deposit; and collecting a gas mixturecomprising the methane.
 2. The method of claim 1, wherein thehydrocarbon-rich deposit is a deposit comprising oil, natural gas, coal,bitumen, tar sands, lignite, peat, carbonaceous shale or sediments richin organic matter.
 3. The method of claim 1, further comprising the stepof characterizing at least one microorganism of the deposit.
 4. Themethod of claim 1, wherein at least one microorganism of the deposit isgenetically modified and reintroduced into the deposit so as to improvenet production of acetate or methane by microorganisms in the deposit.5. The method of claim 1, further comprising the step of introducing atleast one microorganism into the hydrocarbon-rich deposit so as toimprove net acetate or methane production.
 6. The method of claim 1,wherein the at least one environmental parameter characterized comprisestemperature, pH, salinity, sulfate concentration, acetate or otherorganic acid concentration, methane concentration, hydrogenconcentration, water content, metal ion concentration and composition,or hydrocarbon concentration and/or composition.
 7. The method of claim1, wherein the aqueous solution comprises at least one componentselected from the group consisting of complex nutrients, at least onevitamin, at least one metal ion, nitrogen, phosphorus, a trace element,an enzyme, a catalyst and a buffer.
 8. The method of claim 7, whereinthe at least one metal ion is copper, iron, cobalt, nickel ormolybdenum.
 9. The method of claim 1, wherein the aqueous solution isintroduced into the deposit by injection into fractures and/or cleatswithin the deposit.
 10. The method of claim 1, wherein thehydrocarbon-rich deposit is located in an anaerobic geologic formation.11. A method for enhancing microbial production of a gas mixturecomprising methane from an in situ hydrocarbon-rich deposit, the methodcomprising: introducing a catalyst to the in situ deposit, wherein thecatalyst stimulates an increase in production of the methane from the insitu deposit by an amount in excess of the quantity of the catalystintroduced to the in situ deposit; and collecting the gas mixturecomprising the methane.